Figure 1. VHA-5 is at the apical membrane in the excretory canal and the epidermis. (A) Drawing of the V-ATPase complex and C. elegans subunits analyzed in this study. (B) PCR analysis of wild-type,
vha-5(
mc38)/+, and
vha-5(
mc38) animals with primers in
vha-5 showing that
mc38 is a small deletion. (C) Western blot with a VHA-5 antiserum of wild-type C. elegans extracts (lane a) and
vha-5(
mc38) mutants rescued by a
vha-5::gfp transgene (lane b). A 105-kD band is visible in wild-type animals, a 135-kD band in rescued
vha-5(
mc38) mutants. (D) Drawing of a section through the body (left) and the epidermis (right) showing the positions of the images displayed in this and other figures. (E) Distribution of
vha-5::rfp and
vha-8::yfp in rescued
vha-5(
mc38) animals; XY confocal section, apical epidermal surface where the pattern appears as dots (arrowheads; excretory canal, arrows). The V1 E subunit VHA-8 (see A) colocalizes with VHA-5. (F) Immunofluorescence image of a wild-type adult with VHA-5 antiserum; VHA-5 forms dots in the epidermis. (G) Immunogold labeling against VHA-5 (gold beads, arrowheads); VHA-5 localizes mainly to apical membrane stacks of the epidermis (see also Fig. S5 D). Fig. S5 is available at
http://www.jcb.org/cgi/content/full/jcb.200511072/DC1.